Calcitonin receptor (CTR) is a member of a new subfamily of GTP-binding, G protein-coupled receptors (GPCRs) which spans the plasma membrane seven times. Although most GPCRs are expressed in polarized cells, information about polarized distribution and trafficking of this vast and important superfamily of receptors is very limited. I have recently found that the expression of transfected human CTR type 2 (hCTR-2) in Mardin-Darby Canine Kidney (MDCK) cells is highly polarized to the basolateral membrane domain (ratio of 50:1 basolateral to apical). This new model opens an avenue of research to determine how the polarized expression of this receptor is achieved and what are the molecular determinants of this polarization. The proposal is centered on the working hypothesis that information embedded in the sequence and/or structure of the receptor molecule itself is presented to the intracellular sorting machinery as a signal(s) that directs the final fate of CTRs during vesicular traffic (exocytotic/endocytotic pathways, receptor recycling, transcytosis and targeting to lysosomes for degradation). To test this hypothesis, I will pursue the following Specific Aims: 1) To determine the general mechanism (sorting and direct specific targeting versus selective retention) by which CTRs become polarized to the basolateral domain of epithelial cells. 2) To identify the sequences and/or structural elements in CTR that are involved in its domain specific targeting and/or retention to the basolateral domain of polarized epithelial cells. 3) To identify the sequences and/or structural elements in CTR that are involved in CTR internalization, recycling or transcytosis of internalized receptors and CTR degradation. For these aims, mutagenesis of hCTRs will be combined with cell biologic studies. Knowing these mechanisms will not only help to better understand CTR physiology, but also the normal and abnormal function of other GPCRs, e.g. mis-sorting of some mutant opsins causes retinitis pigmentosa. New sorting or targeting signals may be useful in the design of new genes that could make proteins to be expressed at specific locations after delivery by vectors used in gene therapy. Insights derived from this study may lead to the rational design of drugs or therapies that could be used to treat diseases of bone, for example osteoporosis, by affecting calcium homeostasis.